Tumor KRAS Mutation Status Research Articles

Prognostic Factors of Survival in Patients with Peritoneal Metastasis from Colorectal Cancer.

Objectives: The aim of this study was to analyze the prognostic factors of survival in patients with peritoneal metastasis (PM) from colorectal cancer (CRC). The type of relationship between survival and the PM time of detection was used to determine whether it was synchronous with the primary tumor or metachronous. Patients and Methods: Retrospective observational study. It included patients treated for colorectal adenocarcinoma diagnosed between January 2005 and December 2019 who presented PM at the time of diagnosis or during follow-up. Variables, such as sex, age, differentiation grade, positive adenopathy (pN+), tumor size (pT), tumor location, mucinous component, peritoneal carcinomatosis index (PCI), and KRAS mutational status, were analyzed. Results: During the study period, 1882 patients were surgically treated for CRC in our hospital. Of these, 240 patients (12.8%) were included in the study after evidence of PM. The mean age was 67 ± 12 years (range: 32–92 years), and 114 patients were female (47.5%). The mean follow-up was 20 ± 13 months (median 12 months). The Kaplan–Meier survival at 36 months was higher in patients with metachronous PM (24% vs. 8%; p = 0.002), WT-KRAS tumors (31% vs. 15%; p < 0.001), N0 stage (30% vs. 19%; p < 0.001), T3 stage tumors (18% vs. 19% in T4A and 3% in T4B; p > 0.001), and tumors with classic adenocarcinoma histology (18% vs. 8%; p = 0.011). Patients with a PCI of 1–10 showed a likelihood of survival at 36 months of 56%, which was longer than that found in patients with a PCI of 11–20 (8%) or a PCI of >20 (0%) (p < 0.001). In the multiple regression analysis, the factors with an independent prognostic value were: poor grade of differentiation (HR 1.995; 95% CI: 1.294–3.077), KRAS mutation (HR 1.751; 95% CI: 1.188–2.581), PCI 11–20 (HR: 9.935; 95% CI: 5.204–18.966) and PCI > 20 (HR: 4.011; 95% CI: 2.291–7.023). Conclusions: PCI should continue as the as the most useful prognostic indicator in order to assess prognostic estimations as well as therapeutic and surgical decisions, but tumor grade and KRAS mutational status may help in the treatment decision process by providing complementary information. The time of PM detection did not achieve statistical significance in the multiple regression analysis.

Prognosis of Synchronous Colorectal Liver Metastases After Simultaneous Curative-Intent Surgery According to Primary Tumor Location and KRAS Mutational Status.

Tumor location and KRAS mutational status have emerged as prognostic factors of colorectal cancer. We aimed to define the prognostic impact of primary tumor location and KRAS mutational status among synchronous colorectal liver metastases (CRLM) patients who underwent simultaneous curative-intent surgery (SCIS). We compared the clinicopathologic characteristics and long-term outcomes of 227 patients who underwent SCIS for synchronous CRLM, according to tumor location and KRAS mutational status. We cross-classified tumor location and KRAS mutational status and compared survival outcomes between the four resulting patient groups. Forty-one patients (18.1%) had right-sided (RS) tumors and 186 (81.9%) had left-sided (LS) tumors. One-third of tumors (78/227) harbored KRAS mutations. The KRAS mutant-type (KRAS-mt) was more commonly observed among RS tumors than among LS tumors [21/41 (51.2%) vs. 57/186 (30.6%), p = 0.012]. Median follow-up time was 43.4months. Patients with RS tumors had shorter survival times than those with LS tumors [median disease-free survival (DFS): RS, 9.9months vs. LS, 12.1months, p = 0.003; median overall survival (OS): RS, 49.7months vs. LS, 88.8months, p = 0.039]. RS tumors were a negative prognostic factor for DFS [hazard ratio (HR) 1.878, p = 0.001] and OS (HR 1.660, p = 0.060). RS KRAS-mt and LS KRAS wild-type (KRAS-wt) tumors had the worst and best oncological outcomes, respectively. Tumor location has a prognostic impact in patients who underwent SCIS for CRLM, and RS KRAS-mt tumors yielded the worst oncological outcome. These results may allow for more tailored multimodality treatments.

Genomic biomarkers to determine survival in Multicenter Study of RAS mutations (MURAS) in patients with colorectal liver metastases receiving Y90 radioembolization treatment.

e16119 Background: Assessment of the clinical outcomes and prognostic value of genomic mutations in colorectal liver metastases treated with Y90 radioembolization. Methods: Multi-institution retrospective study of patients who underwent Y-90 radioembolization treatment after tumor genotyping was completed for CRLM. Patients treated between 2008 and 2019 were included from 7 institutions within the United States and Europe. Patient demographics, tumor characteristics, pre- and post- treatment regimens, serum laboratory evaluation and overall survival were analyzed between patients with differing histopathologic and genomic status. Tumor genotyping was obtained for KRAS, BRAF, PIK3CA, AKT, MEK, NRAS and MMR genes. Kaplan-Meier survival estimation and multivariate Cox regression were analyzed. Results: 434 patients treated with Y90 radioembolization fulfilled the inclusion criteria. Of the total cohort, 399 patients were available who had sufficient documented tumor profiling data. Average age at diagnosis was found to be 58.8 years for all patients (60.1% male, 39.9% female). Decreased survival post Y-90 treatment was shown in those patients with increased number of documented tumor mutations (n = 0,1 or ≥2 mutations: median OS 9.63 mos vs. 6.2 mos vs. 5.3 mos; p &lt; 0.0001). Additionally, the median survival in patients with mutated BRAF was 5.0 months, as compared to 9.4 months in those patients with wild-type BRAF (p = 0.0009). Primary colon cancer sidedness was also shown to demonstrate significant difference in survival post-Y90 treatment with left sided primaries showing improved median overall survival (left = 7.5 mos vs. right = 6.3 mos; p = 0.04.) Patient demographics including gender, age and race were not shown to be significant in overall survival post-Y90 treatment (p-values &gt; 0.05). Number of tumor mutations (p &lt; 0.0001, HR = 1.69 CI: 1.39-2.05), BRAF status (p = 0.02, HR = 2.6 CI:1.20-4.9), primary sidedness (p = 0.01, HR = 0.65 CI:0.47-0.90), pre-treatment neutrophil-lymphocyte ratio (p = 0.04, HR = 1.42 CI: 1.02-1.98) and KRAS mutation status (p &lt; 0.0001, HR = 1.81 CI: 1.45-2.26) all persisted as significant predictors of survival on multivariate analysis. Conclusions: Number of tumor mutations, BRAF mutation status, primary tumor sidedness, neutrophil-lymphocyte ratio and KRAS mutation status are all shown to be significant prognostic factors in patients with colorectal liver metastases receiving Y90 radioembolization.

Evaluating the Impact of Combination of Nivolumab and Ipilimumab to Liver/Lung SBRT on Local Control for Colorectal and Pancreatic Cancer: A Pooled Analysis of Two Prospective Trials

Immunotherapy (IO) has emerged as an effective treatment option for many metastatic cancers, however, its role in pancreatic or colorectal cancer is limited to those with microsatellite instability. It has been suggested that radiation (RT) and combination IO may work synergistically to increase the immune system’s exposure to tumor antigens, potentiating an abscopal effect. However, the impact of combination IO to stereotactic body radiation therapy (SBRT) on local control of irradiated lesions has not been elucidated. In this study, we combined two prospective phase II single arm studies to compare the impact of SBRT alone (NCT01239381) or with PD-L1/CTLA4 blockade (NCT03104439) on the growth of colorectal and pancreatic liver and lung metastases. Eligible patients had metastatic colorectal or pancreatic adenocarcinoma. Those treated on NCT01239381 received liver SBRT alone (30-50 Gy in 5 fractions), while those on NCT03104439 received combination IO 6 weeks prior to and concurrently with 24 Gy in 3 fractions to liver or lung metastases. Tumor genotyping was performed to assess for KRAS and TP53 mutations. Target lesions were measured on CT scans prior to and following RT. Peak response was measured via modified RECIST criteria as the largest tumor reduction at any time. Wilcoxon Rank Sum tests were performed to evaluate % tumor reduction with regards to combination IO, primary tumor site, metastasis site, and KRAS and TP53 mutation status. There were 50 eligible patients from NCT01239381 and 41 from NCT03104439, of whom 42 and 39 were evaluable, respectively. Of these 81 patients, 52 and 29 had colorectal and pancreatic cancer, respectively. Of those with molecular analysis, 50% (33/66) had a KRAS mutation and 55% (34/61) had a TP53 mutation. Median follow up was 78 days in the IO arm (driven by distant progression) and 160 days in the non-IO arm (P<0.0001). Mean BED in the IO arm was 43.2 Gy vs. 79.6 Gy in the SBRT alone arm (P<0.0001). Mean target size was 3.8 cm in the IO arm, and 5.0 cm in the SBRT alone arm (P=0.12). There was no difference with respect to overall response rate (38.5% vs. 52.4%, P=0.27) and disease control rate (92.3% vs. 92.9%, P>0.99). There was no significant difference in the % tumor reduction between the IO and non-IO arms (-21% vs. -41%, P=0.08). Among the entire cohort, there was no significant difference in response of irradiated lesion with respect to primary tumor type (P= 0.54), KRAS status (P=.67), TP53 status (P=0.10), or metastatic site (P=0.58). Despite receiving approximately half the BED dose compared to liver SBRT alone, patients treated with IO and RT had similar response rates, suggesting a potential synergistic effect in the irradiated lesions. Future studies should look to validate these findings and explore optimal RT dose/fractionation schemas in combination with IO. These results support ongoing randomized studies of IO with RT in locally advanced rectal (NRG GI002) and pancreatic (NCT03563248) cancers.

Metastatic Profile of Colorectal Cancer: Interplay Between Primary Tumor Location and KRAS Status

BackgroundMutant KRAS tumors are purported to metastasize differently than wild-type KRAS tumors. The biological heterogeneity of tumors from different parts of the colon are also reported to affect metastasis. This study aims to characterize the metastatic profile by evaluating these factors in unison. MethodsRetrospective analysis of 899 patients with metastatic colorectal cancers treated from January 2010 to December 2014 was conducted. KRAS mutation status and primary tumors location were correlated with single-site metastasis (liver, lung, and peritoneum) and dual-site metastases (liver-peritoneum, liver-lung, and lung-peritoneum). Patients without KRAS analyses were excluded. ResultsRight-sided tumors had highest frequency of peritoneal metastasis as compared to left-sided or rectal tumors (34.7% versus 15.8% versus 8.8%, P = 0.00) regardless of KRAS status (32.6% versus 38.5%, P = 0.62). Left-sided tumors with wild-type KRAS had greater proportion of liver metastasis (78.6% versus 53.5%, P = 0.00), whereas those with mutant KRAS had greater proportion of lung metastasis (23.3% versus 8.7%, P = 0.02). Rectal tumors with wild-type KRAS tend to spread to the liver (81.4% versus 48.0%, P = 0.00) and not to the peritoneum (2.3% versus 20.0%, P = 0.01). In dual-site metastases, left-sided tumors with wild-type KRAS had more liver-peritoneal metastases (75.0% versus 29.4%, P = 0.00), whereas mutant KRAS had greater lung-liver metastases (64.7% versus 20.8%, P = 0.01). Rectal tumors had the predilection for lung-liver metastases as compared to right-sided and left-sided tumors (92.3% versus 40.0% versus 39.0%, P = 0.00) regardless of KRAS status (100% versus 75%, P = 0.12). ConclusionsOur results may streamline surveillance programs based on primary tumor location and KRAS mutational status.

Metabolic factors and the risk of colorectal cancer by KRAS and BRAF mutation status.

Factors related to energy metabolism and the metabolic syndrome, such as higher body mass index (BMI), blood glucose, or blood lipids, and blood pressure, are associated with an increased risk of colorectal cancer (CRC). However, CRC is a heterogeneous disease, developing through distinct pathways with differences in molecular characteristics and prognosis, and possibly also in risk factors. For subtypes defined by KRAS and BRAF mutation status, BMI is the only metabolic factor previously studied, with inconsistent findings. We investigated whether associations between BMI, blood glucose, blood lipids, and blood pressure and CRC risk differed by tumor KRAS and BRAF mutation status in 117,687 participants from two population-based cohorts within the Northern Sweden Health and Disease Study (NSHDS). Hazard ratios (HRs) for overall CRC and CRC subtypes by metabolic factors were estimated with Cox proportional hazards regression, using multiple imputation to handle missing exposure and tumor data. During a median follow-up of 15.6 years, we acquired 1,250 prospective CRC cases, of which 766 cases had complete baseline and molecular tumor data. Consistent with previous evidence, higher BMI, total cholesterol, triglyceride levels, and blood pressure were associated with an increased risk of overall CRC (HRs per 1 standard deviation increase: 1.07 to 1.12). These associations were similar regardless of CRC subtype by KRAS and BRAF mutation status (all pheterogeneity > 0.05). The same was true for subtypes based on microsatellite instability status. Poor metabolic health may therefore be a universal mechanism for colorectal cancer, acting across multiple developmental pathways.

Abstract 642: Detection of circulating antibodies against KRAS in patients with advanced cancers

Abstract Introduction: KRAS gene plays a major role in tumorigenesis, cell proliferation and survival. Yet, there has been no effective targeted therapy against KRAS mutation-mediated tumors. We hypothesized that KRAS can induce humoral-mediated immune response. Methods: Plasma or serum samples from patients with progressing advanced cancers with or without KRAS mutations were tested for the presence of circulating KRAS antibodies. We used the Maverick Detection System (Genalyte, San Diego, CA), which can perform real-time detection of macromolecules in crude samples using biologically functionalized silicon photonic biosensors lithographically printed on disposable silicon chips. Results: We collected serum or plasma samples from 213 patients with advanced cancers (KRAS-mutant, N=100; KRAS wild-type, N=113) and 50 (23%) were found to have circulating KRAS antibodies. There was no association between KRAS antibodies and tumor KRAS mutation status (21/100, 21% for KRAS-mutant vs. 29/113, 26% for KRAS wild-type; P=0.52). In addition, there was no difference in detection of KRAS antibodies in colorectal cancer (21/89, 24%) compared to other cancers (29/124, 23%; P=1.00). There was no difference in the median survival in patients with KRAS antibodies compared to patients without KRAS antibodies (9.0 months vs. 10.1 months; P= 0.825). Similarly, there was no difference in the median survival according to the presence of circulating KRAS antibodies in subgroups of patients with tumor KRAS mutations (P= 0.96) and without tumor KRAS mutations (P=0.63). On the contrary, the median survival of patients with tumor KRAS mutation was shorter compared to patients with KRAS wild-type (7.2 months vs. 11.5 months; P&amp;lt;0.001). Conclusion: Circulating KRAS antibodies can be detected in 23% of patients with advanced cancers. Biological implications of circulating KRAS antibodies remain to be understood. Citation Format: Ed Kheder, Helen J. Huang, Alice Wu, David S. Hong, Sarina A. Piha-Paul, Daniel D. Karp, Siqing Fu, Vivek Subbiah, Apostolia M. Tsimberidou, Aung Naing, Adi Diab, Milind Javle, Scott Kopetz, Anil K. Sood, Jonathan M. Kurie, Funda Meric-Bernstam, Martin Gleeson, Filip Janku. Detection of circulating antibodies against KRAS in patients with advanced cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 642. doi:10.1158/1538-7445.AM2017-642

Quantitative MR imaging biomarkers of tumor heterogeneity predict prognosis in metastatic colorectal lesions.

e15121 Background: Intra-tumor heterogeneity is an independent determinant of patient survival outcomes in several tumor subtypes. Spatial variations in tumor enhancement on MRI is a macroscopic imaging marker of tumor heterogeneity. The goal of this study is to evaluate the potential role of MRI-based enhancement heterogeneity measures as predictors of survival in patients with metastatic colorectal cancer. Methods: We retrospectively analyzed T1-weighted contrast-enhanced MRI images of metastatic hepatic lesions in 41 patients (mean age 57.2 ±14.2 years) who were diagnosed with stage IV colorectal cancer between 2007 and 2013. Tumor Kras mutation status and patient survival data for up to 95 months was available for all patients. The largest metastatic hepatic lesion was identified by a radiologist and manually segmented. 14 Haralick texture features were extracted from each lesion. Cox proportional hazards regression analysis was used to assess the association between the enhancement heterogeneity measures and patient survival, with adjustment for Kras mutation status as a potential confounder. Backward stepwise feature selection was performed using p &gt; 0.1 (Wald test) to select for statistically significant features. Results:Mean survival time was 39±3.9 months for the study population (51±4.1 months for Kras-wildtype and 37±5.1 for Kras-mutants). 68% of the patients had left-sided colon cancer; 21% had concurrent pulmonary metastases and 0.5% had concurrent brain metastasis. 61%, 27% and 12% of patients were on Flofox, Folfiri or other chemotherapeutic regimen, respectively. Texture matrix homogeneity (HR &gt; 10; p = 0.016), inverse difference moment (HR &lt; 0.1; p = 0.020), entropy (HR &lt; 0.1; p = 0.033) and standard deviation (HR &gt; 10; p = 0.006) exhibited significant independent association with patient survival. With the exception of entropy, these features maintained significant contribution to survival prediction independent of Kras mutation status. Conclusions: MRI-based quantitativeintra-metastasis heterogeneity measures are associated with patient survival and may add information beyond genetic mutation status to optimize prognosis prediction in metastatic colon cancer.

Connecting cancer biology and clinical outcomes to imaging in KRAS mutant and wild-type colorectal cancer liver tumors following selective internal radiation therapy with yttrium-90.

To determine whether pathologic colorectal tumor KRAS mutation status is correlated with progression-free survival (PFS) by imaging after selective internal radiation therapy with Yttrium-90 (SIRT Y90) for metastatic colorectal cancer in the liver (mCRC). This was an IRB approved, HIPAA compliant retrospective cohort study. Consecutive patients with unresectable mCRC with documented KRAS mutation status treated at a single center from 2002 to 2013 with SIRT Y90 were investigated. Treatment response was compared between KRAS wild-type (wt) and mutant (mut) using an anatomic tumor response criteria based on RECIST 1.0. Kaplan-Meier estimation and Cox regression analysis were used to measure progression-free survival (PFS) and to assess independent prognostic factors for PFS. 82 of 186 patients met review criteria. 33 (40.2%) patients were identified as KRAS mut. PFS was longer in KRAS wt (median 166days [95% CI 96-258days]) vs. mut (median 91days [95% CI 79-104days], p=0.002). KRAS mut patients were 1.48 times more likely to progress at first follow-up imaging than wt (95% CI 1.06-2.08, p=0.024). Univariate analysis identified high pre-SIRT Y90 INR, KRAS wt, any use of anti-EGFR therapy, and post-SIRT Y90 chemotherapy as prognostic factors for longer PFS. In multivariate analysis, only KRAS wt was an independent prognostic factor for longer PFS (RR: 1.80 [95% CI 1.08-2.99], p=0.024). Longer PFS is associated with KRAS wt vs. mut following SIRT Y90.

Detection and Analysis of EGFR and KRAS Mutation with Lung Adenocarcinoma

背景与目的表皮生长因子受体(epidermal growth factor receptor, EGFR)突变和KRAS基因突变是非小细胞肺癌(non-small cell lung cancer, NSCLC)靶向治疗的重要分子标志，与临床治疗疗效密切相关。分析肺腺癌EGFR和KRAS基因突变与临床特征的关系。方法收集初治肺腺癌患者395例，有可供检测的肿瘤组织标本。利用突变富集液相芯片法进行EGFR和KRAS基因突变检测。结果395例肺腺癌中，EGFR基因突变192例(48.9%)，KRAS基因突变29例(7.8%)，EGFR和KRAS基因同时发生突变1例(0.3%)。女性和不吸烟患者EGFR基因突变率高于男性和吸烟患者(62.0% vs 37.1%, 61.9% vs 30.3%)，差异具有统计学意义(P＜0.001, P＜0.001)；不同年龄、分期及病理取材标本之间差异均无统计学意义(P＞0.05)。KRAS基因突变在EGFR基因野生型患者中发生率(13.5%, 27/200)明显高于EGFR基因突变患者(1.0%, 2/192)，差异具有统计学意义(P＜0.001)。结论肺腺癌EGFR基因突变在女性和不吸烟患者中较高，KRAS基因突变只在EGFR基因野生型患者中较高。在使用TKI靶向治疗药物之前，应同时检测EGFR基因和KRAS基因的突变情况。

Panitumumab Use in Metastatic Colorectal Cancer and Patterns of KRAS Testing: Results from a Europe-Wide Physician Survey and Medical Records Review.

BackgroundFrom 2008–2013, the European indication for panitumumab required that patients’ tumor KRAS exon 2 mutation status was known prior to starting treatment. To evaluate physician awareness of panitumumab prescribing information and how physicians prescribe panitumumab in patients with metastatic colorectal cancer (mCRC), two European multi-country, cross-sectional, observational studies were initiated in 2012: a physician survey and a medical records review. The first two out of three planned rounds for each study are reported.MethodsThe primary objective in the physician survey was to estimate the prevalence of KRAS testing, and in the medical records review, it was to evaluate the effect of test results on patterns of panitumumab use. The medical records review study also included a pathologists’ survey.ResultsIn the physician survey, nearly all oncologists (299/301) were aware of the correct panitumumab indication and the need to test patients’ tumor KRAS status before treatment with panitumumab. Nearly all oncologists (283/301) had in the past 6 months of clinical practice administered panitumumab correctly to mCRC patients with wild-type KRAS status. In the medical records review, 97.5% of participating oncologists (77/79) conducted a KRAS test for all of their patients prior to prescribing panitumumab. Four patients (1.3%) did not have tumor KRAS mutation status tested prior to starting panitumumab treatment. Approximately one-quarter of patients (85/306) were treated with panitumumab and concurrent oxaliplatin-containing chemotherapy; of these, 83/85 had confirmed wild-type KRAS status prior to starting panitumumab treatment. All 56 referred laboratories that participated used a Conformité Européenne-marked or otherwise validated KRAS detection method, and nearly all (55/56) participated in a quality assurance scheme.ConclusionsThere was a high level of knowledge amongst oncologists around panitumumab prescribing information and the need to test and confirm patients’ tumors as being wild-type KRAS prior to treatment with panitumumab, with or without concurrent oxaliplatin-containing therapy.

Distinct gene expression profiles of proximal and distal colorectal cancer: implications for cytotoxic and targeted therapy

Colorectal cancer (CRC) is a heterogeneous disease with genetic profiles and clinical outcomes dependent on the anatomic location of the primary tumor. How location impacts the molecular makeup of a tumor and how prognostic and predictive biomarkers differ between proximal versus distal colon cancers is not well established. We investigated the associations between tumor location, KRAS and BRAF mutation status, and the mRNA expression of proteins involved in major signaling pathways, including tumor growth (EGFR), angiogenesis (VEGFR2), DNA repair (ERCC1) and fluoropyrimidine metabolism (TS). FFPE tumor specimens from 431 advanced CRC patients were analyzed. The presence of 7 different KRAS base substitutions and the BRAF V600E mutation was determined. ERCC1, TS, EGFR and VEGFR2 mRNA expression levels were detected by RT-PCR. BRAF mutations were significantly more common in the proximal colon (p<0.001), whereas KRAS mutations occurred at similar frequencies throughout the colorectum. Rectal cancers had significantly higher ERCC1 and VEGFR2 mRNA levels compared to distal and proximal colon tumors (p=0.001), and increased TS levels compared to distal colon cancers (p=0.02). Mutant KRAS status was associated with lower ERCC1, TS, EGFR, and VEGFR2 gene expression in multivariate analysis. In a subgroup analysis, this association remained significant for all genes in the proximal colon and for VEGFR2 expression in rectal cancers. The mRNA expression patterns of predictive and prognostic biomarkers as well as associations with KRAS and BRAF mutation status depend on primary tumor location. Prospective studies are warranted to confirm these findings and determine the underlying mechanisms.

Abstract 5607: BRAF and KRAS mutation testing in plasma cell-free DNA with ICE COLD-PCR in patients with advanced cancers

Abstract Background: Cell-free (cf) DNA in the plasma of cancer patients offers an easily obtainable, low-risk, inexpensive, and repeatedly available source of biologic material for mutation analysis and monitoring of molecular changes throughout cancer therapy. Methods: DNA in plasma from patients with advanced cancers who progressed on systemic therapy was tested for BRAF V600 and KRAS G12 and G13 mutations using the ICE COLD-PCR platform. ICE COLD-PCR, “Improved and Complete Enrichment COamplification at Lower Denaturation,” selectively amplifies mutant DNA by exploiting differences in denaturation temperatures between mutant DNA duplexes and normal “wild-type” DNA duplexes. KRAS Exon 2 and BRAF Exon 15 ICE COLD-PCR was performed on plasma samples. Amplicons were analyzed using Sanger sequencing and results were compared to the mutation status of archival primary or metastatic tumor tissue as determined in a CLIA-certified laboratory during routine clinical care. Results: Plasma samples from 77 patients with advanced cancers and known tumor tissue BRAF and/or KRAS mutation status (colorectal cancer, n=38; melanoma, n=17; non-small cell lung cancer, n=7; other cancers, n=15) were obtained before treatment and, if possible, sequentially during therapy and tested for BRAF (42), KRAS (34) or BRAF and KRAS (1) mutations in cfDNA. BRAF mutations were detected in 93% (40/43) of archival tumor samples compared to 70% (30/43) of plasma cfDNA samples (agreement 77%). In addition, 20 patients treated with systemic therapy had serial plasma samples collected and the change in relative abundance of BRAF-mutant compared to wild-type cfDNA corresponded with the clinical course of 15 patients and was discrepant for 1 patient; in 5 patients no BRAF mutated cfDNA was detected at any time point. KRAS mutations were detected in 83% (29/35) of archival tumor samples compared to 74% (26/35) of plasma cfDNA samples (agreement 80%). In addition, 12 patients treated with systemic therapy had serial plasma collected and the change in relative abundance of KRAS-mutant compared to wild-type cfDNA corresponded with clinical course in 10 patients; in 2 patients no KRAS mutated cfDNA was detected at any time point. Conclusions: Detection of BRAF and KRAS mutations in cfDNA can provide a fast and noninvasive alternative to mutation testing in tumor tissue with a potential to be used for monitoring response to cancer therapy. Citation Format: Filip Janku, Ben Legendre, Katherine Richardson, Gerald S. Falchook, Aung Naing, Veronica R. Holley, Siqing Fu, David S. Hong, Sarina A. Piha-Paul, Jennifer J. Wheler, Ralph G. Zinner, Vivek Subbiah, Apostolia M. Tsimberidou, Daniel D. Karp, Vanda M. Stepanek, Goran Cabrilo, Rajyalakshmi Luthra, Funda Meric-Bernstam, Agop Y. Bedikian, Bryan K. Kee, Cathy Eng, Michael J. Overman, Kevin B. Kim, Amy Kruempel, Jaclyn Pope, Courtney Cubrich, Grant Wu, Marcia Lewis, Razelle Kurzrock. BRAF and KRAS mutation testing in plasma cell-free DNA with ICE COLD-PCR in patients with advanced cancers. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5607. doi:10.1158/1538-7445.AM2014-5607

Outcomes by Tumor Histology and KRAS Mutation Status After Lung Stereotactic Body Radiation Therapy for Early-Stage Non–Small-Cell Lung Cancer

BackgroundWe analyzed outcomes after lung stereotactic body radiotherapy (SBRT) for early-stage non–small cell lung-carcinoma (NSCLC) by histology and KRAS genotype. Patients and MethodsWe included 75 patients with 79 peripheral tumors treated with SBRT (18 Gy × 3 or 10 to 12 Gy × 5) at our institution from 2009 to 2012. Genotyping for KRAS mutations was performed in 10 patients. Outcomes were analyzed by the Kaplan-Meier method/Cox regression, or cumulative incidence method/Fine-Gray analysis. ResultsThe median patient age was 74 (range, 46 to 93) years, and Eastern Cooperative Oncology Group performance status was 0 to 1 in 63%. Tumor histology included adenocarcinoma (44%), squamous cell carcinoma (25%), and NSCLC (18%). Most tumors were T1a (54%). Seven patients had KRAS-mutant tumors (9%). With a median follow-up of 18.8 months among survivors, the 1-year estimate of overall survival was 88%, cancer-specific survival (CSS) 92%, primary tumor control 94%, and freedom from recurrence (FFR) 67%. In patients with KRAS-mutant tumors, there was a significantly lower tumor control (67% vs. 96%; P = .04), FFR (48% vs. 69%; P = .03), and CSS (75% vs. 93%; P = .05). On multivariable analysis, histology was not associated with outcomes, but KRAS mutation (hazard ratio, 10.3; 95% confidence interval, 2.3-45.6; P = .0022) was associated with decreased CSS after adjusting for age. ConclusionIn this SBRT series, histology was not associated with outcomes, but KRAS mutation was associated with lower FFR on univariable analysis and decreased CSS on multivariable analysis. Because of the small sample size, these hypothesis-generating results need to be studied in larger data sets.

149IN - Ttd Contribution to the Precision Oncology

ABSTRACT The TTD Group is a Spanish Cooperative Group with 28 years of history investigating and developing protocols in the digestive tumor field to improve healthcare quality. In the last decade, it research has been focused on the identification of biomarkers to improve and predict treatment outcomes. The MACRO (Maintenance in Colorectal Cancer) study was a phase III study to assess maintenance therapy with single-agent bevacizumab (B) versus B plus chemotherapy (QT) in patients (p) with metastatic colorectal cancer (mCRC). A post hoc analysis of the trial investigated the prognostic value of the circulating tumor cell (CTC) count and KRAS mutation status on patient outcomes and showed that both factors were independent prognostic factors for outcomes. Based on these results, the ongoing VISNU study will screen 750 p previously untreated with mCRC according to CTC count and gene status (RAS, BRAF y PI3K). In VISNU-1 trial (NCT01640405), 350 p with ≥ 3 CTC, will be randomized to receive FOLFOX-6 + B or FOLFOXIRI + B. In VISNU-2 trial (NCT01640444), 240 p with Disclosure: E. Aranda Aguilar: Honoraria or consultation fees: Roche and Merck.

Associations between patient (pt) colorectal cancer (CRC) tumor KRAS and BRAF mutation (mut) status and overall survival (OS).

473 Background: Evolving information is emerging regarding patient tumor molecular data and associations with patient clinical characteristics and survival outcomes; we sought to evaluate pt clinical data with CRC tumor mut status. Methods: Since January 2008 tumor pts with stage IV CRC (principally primary tumors) were analyzed for KRAS (codon 12, 13, and 61) mut status as standard-of-care. A subset of pts tumors underwent BRAF and KRAS codon 164 mut testing. Results: 1,891 pts with CRC whose tumors underwent molecular mut analysis were identified from the clinical record. There was no statistically significant difference in pt overall survival from time of diagnosis for stages I/II (data not shown) and III; for stage IV pts there was an inferior OS associated with BRAF mut tumors (see Table). Tumor BRAF mut was significantly detected among ever smokers (10.2%) as compared to never smokers (5.4%), p &lt; 0.001; meanwhile, tumor KRAS mut and wild type distribution were similar between ever smokers and never smokers. KRAS mut was significantly observed in CRC patients with distant lymph node, hepatic and lung metastasis, p &lt; 0.001. Type 2 diabetes, intake of metformin or ACE inhibitors, were not significantly correlated with tumor KRAS or BRAF mut status. Conclusions: Pts with stage IV CRC BRAF mut tumors have a higher likelihood of prior or current smoking history in addition to having an adverse survival prognosis; pts with stage IV KRAS mut tumors have an increased association with distant lymph node, hepatic, and lung metastases as compared to KRAS wild type and BRAF mut tumors. Studies are ongoing to determine how tumor molecular characteristics may help to predict clinical outcomes and provide a useful to guide pt care. [Table: see text]

Tumor Heterogeneity Revealed byKRAS,BRAF, andPIK3CAPyrosequencing:KRASandPIK3CAIntratumor Mutation Profile Differences and Their Therapeutic Implications

Current clinical problems in colorectal cancer (CRC) diagnostics and therapeutics include the disease complexity, tumor heterogeneity, and resistance to targeted therapeutics. In the present study, we examined 171 CRC adenocarcinomas from Greek patients undergoing surgery for CRC to determine the frequency of KRAS, BRAF, and PIK3CA point mutations from different areas of tumors in heterogeneous specimens. Ninety two out of 171 (53.8%) patients were found to bear a KRAS mutation in codons 12/13. Of the 126 mutations found, 57.9% (73/126) were c.38G>A mutations (p.G13D) and 22.2% (28/126) were c.35G>T (p.G12V). Remarkably, RAS mutations in both codons 12 and 13 were recorded in the same tumor by pyrosequencing. Moreover, differences in KRAS mutations between tumor center and periphery revealed tumor heterogeneity in 50.7% of the specimens. BRAF c.1799T>A (V600E) mutations were moderately detected in 4/171 (2.3%) specimens, whereas most PIK3CA mutations were revealed by pyrosequencing 6/171 (3.5%). Remarkable tumor heterogeneity is revealed, where double mutations of KRAS in the same tumor and different KRAS mutation status between tumor core and margin are detected with high frequency. It is expected that these findings will have a major impact in cancer diagnosis and personalized therapies.

FOLFOX4 With Cetuximab vs. UFOX With Cetuximab as First-Line Therapy in Metastatic Colorectal Cancer: The Randomized Phase II FUTURE Study

IntroductionThe purpose of this study was to assess the efficacy and safety of FOLFOX4, comprising infusional 5-fluorouracil (5-FU)/leucovorin (LV) and oxaliplatin, with cetuximab compared with UFOX, comprising UFT, an oral prodrug of 5-FU, LV, and oxaliplatin, with cetuximab as first-line treatment for mCRC. Patients and MethodsPatients, unselected by tumor KRAS status, were randomized 1:1 to FOLFOX4 with cetuximab or UFOX with cetuximab. Treatment was continued until disease progression or unacceptable toxicity. The primary end point, assessed in the intention-to-treat population, was progression-free survival (PFS). Secondary end points included tumor response, overall survival, and safety. Outcome according to KRAS mutation status was investigated. ResultsRecruitment was curtailed at 302 patients after reporting of the importance of tumor KRAS mutation status for cetuximab activity. Baseline characteristics were balanced between treatment groups. PFS was significantly longer in the FOLFOX4 with cetuximab group compared with UFOX with cetuximab group (median 8.2 vs. 6.6 months; hazard ratio, 0.68; 95% confidence interval [CI], 0.52-0.89; P = .0048). The response rate was also significantly greater in the FOLFOX4 with cetuximab group (51.3% vs. 37.5%, respectively; odds ratio, 1.76; 95% CI, 1.11-2.78; P = .0160), although overall survival was comparable. In the KRAS wild type subgroup, efficacy outcomes were similar to those in the intention-to-treat population. Side effect profiles were manageable and consistent with expectations. ConclusionIn the first-line treatment of mCRC, UFOX with cetuximab had an acceptable safety profile but inferior activity compared with FOLFOX4 with cetuximab in relation to PFS and response. The regimens were comparable with regard to overall survival.

EGFR ligands as pharmacodynamic biomarkers in metastatic colorectal cancer patients treated with cetuximab and irinotecan

This study was conducted to describe the modulation of plasma epidermal growth factor receptor (EGFR) ligands in EGFR-positive metastatic colorectal cancer (mCRC) patients during treatment with cetuximab and irinotecan and to explore the clinical implication of plasma levels' variations as potential biomarkers of benefit. Plasma amphiregulin (AR), epidermal growth factor (EGF), transforming growth factor-α, and heparin binding-EGF were assessed by ELISA in 45 chemorefractory mCRC patients, treated with cetuximab and irinotecan. Plasma levels were measured before and 1h after the first administration of cetuximab, before and 1h after the second administration, and before the third and the fifth cycles. KRAS and BRAF mutational status were determined. EGFR ligands' levels were differently modulated according to tumor KRAS and BRAF mutational status. In KRAS wild-type patients (n = 34), AR and EGF early increased and higher increases were significantly associated with worse clinical outcome. By adopting a specific cut-off value, patients with higher levels of AR 1h after the first administration had significantly worse response rate, progression free survival, and overall survival. This hypothesis-generating study shows that EGFR ligands are significantly modulated by cetuximab plus irinotecan according to KRAS and BRAF mutational status, and they warrant further investigation as pharmacodynamic markers of resistance to anti-EGFRs.

Colorectal carcinomas with KRAS mutation are associated with distinctive morphological and molecular features

KRAS-mutated carcinomas comprise 35–40% of all colorectal carcinomas but little is known about their characteristics. The aim of this study was to examine the pathological and molecular features of KRAS-mutated colorectal carcinomas and to compare them with other carcinoma subgroups. KRAS mutation testing was performed in 776 incident tumors from the Melbourne Collaborative Cohort Study. O6-methylguanine DNA methyltransferase (MGMT) status was assessed using both immunohistochemistry and MethyLight techniques. Microsatellite instability (MSI) phenotype and BRAF V600E mutation status were derived from earlier studies. Mutation in KRAS codon 12 or codon 13 was present in 28% of colorectal carcinomas. Compared with KRAS wild-type carcinomas, KRAS-mutated carcinomas were more frequently observed in contiguity with a residual polyp (38 vs 21%; P<0.001), demonstrated mucinous differentiation (46 vs 31%; P=0.001) and were associated with different MSI status (P<0.001) and with MGMT methylation (47 vs 21%; P=0.001). Compared with tumors demonstrating neither BRAF nor KRAS mutation, KRAS-mutated carcinomas showed more frequent location in the proximal colon (41 vs 27%; P=0.001), mucinous differentiation (46 vs 25%; P<0.001), presence of a contiguous polyp (38 vs 22%; P<0.001), MGMT methylation (47 vs 26%; P=0.01) and loss of MGMT immunohistochemical expression (27 vs 19%; P=0.02). KRAS-mutated carcinomas were distributed in a bimodal pattern along the proximal–distal axis of the colorectum. Compared with male subjects, female subjects were more likely to have KRAS-mutated carcinoma in the transverse colon and descending colon (39 vs 15%; P=0.02). No difference in overall survival was observed in patients according to their tumor KRAS mutation status. In summary, KRAS-mutated carcinomas frequently develop in contiguity with a residual polyp and show molecular features distinct from other colorectal carcinomas, in particular from tumors with neither BRAF nor KRAS mutation.